The site of macromolecular restriction in the renal glomerulus will be defined by several new morphological approaches for electron microscopy. First, the permeability barrier in the mouse glomerulus will be probed with three new series of ultrastructural tracers with molecular weights which span the cut-off size for the filter as defined by serum albumin. These tracers will consist of (a) horseradish peroxidase (HRP) conjugated to IgG immunoglobulin and its Fab and (F(ab')2 fragments, (b) synthetic oligomers of HRP, and (c) polyvinylpyrrolidone (PVP) fractions of graded molecular weights. HRP-derived tracers will be visualized histochemically with hydrogen peroxide and 3, 3'-diaminobenzidine. PVP fractions will be detected by a fixation technique which employs tannic acid. With each series of tracers, the relative barrier functions of the basement membrane (BM) and slit diaphragm (SD) will be ascertained. As a second approach, the morphology of the glomerular wall in the bullfrog will be described as an example where the BM exhibits an altered ultrastructure which may influence its filtration properties. Horse spleen ferritin will be used as an ultrastructural tracer to assess the permeability of the BM and SD to proteins in this species. Finally, the comparative ultrastructure of the SD in several other animal species will be investigated to determine whether the characteristically small, rectangular pores found in the SD of frogs and mammals are also present in the SD of other species whose sera contain albumin.